Using human ESCs as a genetic model for PNH and other blood diseases

描述(申请人提供)：阵发性睡眠性血红蛋白尿(PNH)是一种造血干细胞(HSCs)获得PIG-A基因突变的克隆性疾病。在几乎所有检测的PNH专利中都发现了克隆的PIG-A突变，导致在受影响的HSC和所有衍生的造血后代中缺乏所有GPI锚定蛋白(GPI-AP)。尽管PNH的生化和分子机制已被阐明，但PNH患者中PIG-A突变克隆性优势的机制以及PNH与其他骨髓衰竭疾病如再生障碍性贫血(AA)和骨髓发育不良综合征(MDS)的密切关系仍不清楚。在临床症状出现之前的早期阶段监测患者的PIG-A突变是不可能的。目前，在健康志愿者的人类HSCs中创建PIG-A突变是不可行的，并且现有的PIG-A空小鼠缺乏GPI-AP。血细胞没有复制在患者中看到的PNH症状。这项研究的唯一目标是了解GPI-APs在骨髓衰竭综合征和造血中的作用。针对PA-05-013，一种基于人类胚胎干细胞[NIH批准，WA01(H1)]的前瞻性实验系统被提出，以研究PIG-A/GPI-AP缺陷对人造血细胞的影响。NIH批准的含有诱导PIG-A突变且GPI-AP缺失的人ESC将被使用，并将使用最近开发的人类ESC启动的造血系统来检测PIG-A/GPI-AP缺乏症的影响。该项目的成功完成也可能为研究正常和异常的人类造血和HSCs提供一种新的遗传模型。对人类造血和造血干细胞中PIG-A/GPI-AP缺陷的深入了解将反过来帮助我们改进和开发PNH和其他相关血液疾病的新治疗方法，如AA和MDS。

DESCRIPTION (provided by applicant): Paroxysmal Nocturnal Hemoglobinuria (PNH) is a clonal disorder of hematopoietic stem cells (HSCs) acquiring mutations in the PIG-A gene. Clonal PIG-A mutations are found in nearly all PNH patents tested resulting in the lack of all GPI-anchored proteins (GPI-APs) in affected HSCs and all the derived hematopoietic progeny. Despite evidences that the biochemical and molecular mechanisms for PNH have been brilliantly elucidated, mechanisms of PIG-A mutant clonal dominance in PNH patients and the close relationship of PNH to other marrow failure diseases such as aplastic anemia (AA) and myelodysplasia syndrome (MDS) are still unknown. Monitoring the PIG-A mutations in patients is impossible at early stage before the onset of clinical features. Creating a PIG-A mutation in human HSCs from healthy volunteers is not feasible currently and existing Pig-a null mice lacking GPI-APs in. blood cells have not replicated the PNH symptoms seen in patients. The lone term objective of this research is understanding effects of GPI-APs in bone marrow failure syndrome and hematopoesis. In response to PA-05-013, a human ESC-based [NIH-approved, WA01(H1)], prospective experimental system is proposed here to investigate effects of the PIG-A/GPI-AP deficiency in human hematopoietic cells. The NIH-approved human ESC that contain an induced PIG-A mutation and are GPI-APs deficient will be used, and effects of PIG-A/GPI-AP deficiency using human ESC-initiated hematopoiesis systems recently developed will be examined. The successful completion of this project may also provide a novel genetic model to investigate normal and abnormal human hematopoiesis and HSCs. Improved understanding of PIG-A/GPI-AP deficiency in human hematopoiesis and HSCs will in turn help us to improve and develop new treatments for PNH and other related blood diseases such as AA & MDS.